Neisseria gonorrhoeae (gonococcus, GC) not only causes uncomplicated gonorrhea but also complications like pelvic inflammatory disease and disseminated gonococcal infection. Pilus is an important pathogenicity factor of GC and is a polymer of the glycoprotein pilin. Glycans of pilin were proposed to play important role in GC pathogenesis. We have cloned a pilin galactosyl transferase (pgtA), that adds an alpha galactosyl. In most complicated disease strains, pgtA contains a poly-G tract like the other phase-variable genes that are frequently turned on and off to presumably provide GC with selective advantage in different niches of human body. However, in uncomplicated gonorrheal isolates, this poly- G is generally not present in pgtA. Hence, we hypothesize that the poly-G mediated high-frequency phase-variation of pgtA plays an important role in conversion of uncomplicated gonorrhea to more complex systemic diseases. In addition, our High pH Anion Exchange Chromatographic analysis of GC pilin glycans suggests presence of sugars that were not reported before. These observations lead to the hypothesis that GC pilin glycans are considerably more complex than the current models and some of these glycans can be associated with certain disease phenotypes or with particular sites of the human body. To test the stated hypotheses, the following specific aims are proposed: 1) Characterization of GC pilin glycans from selective strains and pilus glycosylation mutants and identification of novel pilin glycosylation genes; and 2) Examination of the role of pilin glycans in GC pathogenesis. These studies will further elucidate the mechanism of gonococcal infections. Additionally, glycosylation of many surface proteins, including pilins, of several pathogenic bacteria is being reported. Therefore, the study of the role of pilin glycans in GC pathogenesis is likely to provide important leads into the mechanisms of pathogenesis of other bacteria as well.